Radiation exposure during cancer treatment can cause significant cardiac morbidity and mortality both acutely and years after the initial exposure. These short and long-term complications are increasingly important as new therapies for cancer are developed and survival improves. Radiation can directly damage the myocardium, cause coronary artery and valvular disease, disrupt cardiac innervation and precipitate myocardial fibrosis; this can subsequently lead to systolic and diastolic dysfunction along with atrial and ventricular arrhythmias. In addition, radiation therapy is often given concomitantly with chemotherapy and it is difficult to sort out the direct effects of radiation. The detailed mechanisms underlying the susceptibility to tachy- and bradyarrhythmias following radiation therapy have not been defined. We now show that short and long-term total body and cardiac targeted irradiation in mice leads to oxidative stress, conduction system disease, serum miR-34a upregulation, arrhythmias and increased mortality which were prevented, in part, by treatment with the water soluble oxetanyl sulfoxide compound MMS350 that was developed as a radiation mitigator at the University of Pittsburgh. We also show that Nitric Oxide Synthase 1 knockout (Nos1-/-) mice develop more severe conduction disease associated with increased mortality, that Nos1 genotype alters the effects of radiation on mitochondrial function and reactive oxygen species (ROS) metabolic pathways, that Nos1 genotype alters the metabolic response to MMS350 following irradiation. Our primary hypothesis is that NOS1 and microRNAs play heretofore unrecognized roles in radiation-induced damage to the heart and cardiac conduction system by mitigating changes in oxidative stress and mitochondrial dysfunction that lead to electrophysiological remodeling and arrhythmias, and that MMS350 can mitigate the arrhythmogenic phenotype by protecting cellular metabolism and energetics. To test these hypotheses, we will use total body and cardiac targeted irradiation of wild type and genetically engineered mice to determine the mechanisms by which conduction disease and arrhythmias result from damage to cardiac myocytes, vascular smooth muscle and endothelial cells and/or the autonomic nervous system, and to determine the mechanisms by which NOS1, MMS350 and microRNAs alter the damage. These studies will define the role of Nos1 and microRNAs in cardiac conduction and arrhythmias following radiation exposure and test pharmacological therapies that may mitigate this potentially lethal side effect of a life-saving cancer therapies.